This application is a 371 of PCVEP99/03250 filed May 7, 1999.
The present invention relates to the field of acylation of amino alcohols.
Ceramides are the main lipid component in the upper layer of the skin, the stratum corneum. This skin layer has an important barrier function in that external compounds generally are kept out whereas loss of moisture from the skin is limited. Ceramides are applied in cosmetics for instance because of their moisture retaining effects on the skin.
Typically, ceramides for use in cosmetics are obtained either from a natural source, via chemical synthesis routes or via a combined fermentative and chemical synthesis process. The latter routes are preferred due to the fact that potentially harmful infectious agents may be present in mammalian sources.
Various methods to synthesize ceramides are known in the art. The methods most frequently used involve a sphingoid base and a suitable fatty acid component as the starting materials. The fatty acid component is thereby coupled to the amino group of the sphingoid base via an amide linkage.
WO 93/20038 describes the acylation of amino alcohols whereby the fatty acid is coupled to the amino group as a mixed anhydride. It is essential for this reaction using a mixed anhydride that it occurs under essentially non-aqueous conditions.
Philippe et al. (Int. J. Cosm. Sci. 17, 133-146, 1995) describe an acylation method wherein the fatty acid is coupled to the amine as an acid halogenide, using tetrahydrofuran (THF) as the solvent and triethylamine as the organic base. In this method, the coupling reaction also occurs in a non-aqueous environment.
The requirement for non-aqueous conditions is an important disadvantage in those cases that one or both starting compounds are delivered as a water-containing material. For instance, in the case that the sphingoid base is contained within a wet crystal cake originating from a microbial fermentation process.
A further disadvantage of the above desribed methods is that they only result in a moderate product yield.
Still another acylation method is known applying a solvent system wherein THF is mixed with an equal volume of a 50% NaAc solution (see EP 212400). Although water is present in this system, the additional presence of a high salt concentration is required. A high salt concentration is undesired in that it is expensive and increases the waste load.
It is therefore desirable to be able to apply an acylation process wherein it is not required that the sphingoid base reactant is essentially water-free, which additionally does not require a high salt concentration and which gives a higher product yield than the currently known processes.
The present invention discloses a process for the acylation of an amino alcohol using a fatty acid being in the form of an acid halogenide, wherein the coupling occurs in an organic solvent in the additional presence of water.
The fact that water is present in the reaction mixture has several important advantages. For instance, it is not necessary to dry the amino alcohol reactant before applying the same in the acylation process. This is especially advantageous when the amino alcohol, e.g. a sphingoid base, is obtained via microbial fermentation or as a product from a reaction occurring in an aqueous environment. In addition, the pH of the process can be advantageously controlled using a simple mineral base, such as NaOH, instead of a potentially hazardous organic base.
The process of the invention further neither requires the use of a high salt concentration, as is the case for the process disclosed in EP 212400, nor the use of stoichiometric amounts of auxiliary chemicals, as is the case for the process disclosed in WO93/20038. A further important advantage is that the acylated reaction product is recovered from the organic phase by simply washing the organic phase with water followed by azeotropic removal of water and crystallization. Unexpectedly, the process of the invention provides an acylated amino alcohol in a yield which is considerably higher than the yields obtained in WO 93/20038 or by Philippe et al. In addition, the product has a high purity, i.e. does not contain undesired byproducts, since O-acylation occurs only to a very minor extent.
In the process of the invention, an amino alcohol is acylated with an organic acid of formula RCOOH, the organic acid being in the form of an acid halogenide, said process comprising the steps of:
suspending or dissolving the amino alcohol, or a salt thereof, in an organic solvent in the presence of 0.01 to 10 volumes of water to one volume of organic solvent,
adding the organic acid halogenide as a pure compound or as a solution or suspension in the organic solvent used to suspend the amino alcohol, while keeping the pH at a value of about 5 to 12,
stirring the resulting mixture until the amino alcohol is converted to the N-acylated compound,
recovering the N-acylated aminoalcohol from the organic phase.
In the organic acid of formula RCOOH, R is hydrogen, an optionally unsaturated, optionally substituted, optionally one or more heteroatoms containing straight chain or branched alkyl group having up to 55 carbon atoms; an optionally unsaturated, optionally substituted, optionally one or more heteroatoms containing C5-8 cycloalkyl group; an optionally substituted aryl or heteroaryl group; or an optionally substituted benzyl group.
In a preferred embodiment of the invention, the alkyl group is optionally interrupted by an oxygen atom or by an internal ester group. In another preferred embodiment, the alkyl group has 1 to 50 carbon atoms, more preferably 10 to 50 carbon atoms, most preferably 15 to 45 carbon atoms.
A preferred substituent of the above defined groups is a hydroxyl group, especially an xcex1-hydroxyl group.
In a preferred embodiment of the invention, the organic acid of formula RCOOH is hexanoic acid, octanoic acid, stearic acid, oleic acid, linoleic acid, 27-stearoyloxy-heptacosanoic acid, 27-linoleoyloxy-heptacosanoic acid, xcex1-hydroxy-stearic acid, lactic acid, retinoic acid, salicylic acid or ferulic acid.
Protecting groups for the optional hydroxyl groups are well known in the art and may be selected from appropriate groups as disclosed in Greene, T. (1981) Protective Groups in Organic Synthesis (John Wiley and Sons; New York). In one embodiment of the invention, a hydroxyl group is protected as an acetyl ester or a methoxy methyl ether.
In another embodiment of the invention, a hydroxyl group is provided by first coupling an acid halogenide containing a halogen group at the position corresponding to the future hydroxyl group and subsequently converting the halogen group, i.e. after coupling, to an oxygen function, for instance an acetoxy group. Conversion of the halogen group into an oxygen function and subsequent conversion of the oxygen function into a hydroxyl group are conveniently performed using commonly known methods. Preferably, the halogen group is a bromine group.
The present invention also envisages the option to use a mixture of related organic acids, for instance a mixture of fatty acids having an alkyl group of different chain lengths and/or different extent of unsaturation.
In another preferred embodiment of the invention, the amino alcohol is a sphingoid base of formula
Rxe2x80x2xe2x80x94Axe2x80x94CH(ORxe2x80x2xe2x80x3)xe2x80x94C(NH2)xe2x80x94CH2xe2x80x94ORxe2x80x3
or a salt thereof, wherein:
Rxe2x80x2 is a straight chain or branched alkyl group having 10 to 22 carbon atoms which may optionally contain one or more double bonds and/or may optionally be substituted, preferably with one or more hydroxyl groups, preferably is a straight chain alkyl group having 12 to 18 carbon atoms, more preferably is a straight chain alkyl group having 13 carbon atoms, and
Rxe2x80x3 is hydrogen or a carbohydrate, such as a hexose or pentose moiety (optionally linked to further carbohydrate moieties), preferably hydrogen or a glucose or galactose moiety,
A is CH2xe2x80x94CH2, CHxe2x95x90CH or CH2xe2x80x94C(H)ORxe2x80x2xe2x80x3, and Rxe2x80x2xe2x80x3 is hydrogen or an acyl group of 1 to 3 carbon atoms, preferably is hydrogen or an acetyl group.
The sphingoid base which is used in the method of the invention preferably is a sphingosine, a sphinganine or a phytosphingosine. More preferably, the sphingoid base is phytosphingosine.
In a especially preferred embodiment of the invention, phytosphingosine is obtained via deacetylation of tetraacetylphytosphingosine derived from a microbial fermentation, e.g. of the yeast Pichia ciferri. It is also possible to employ an acetylphytosphingosine obtained by partial deacetylation of tetraacetylphytosphingosine. The present invention advantageously allows that phytosphingosine or a partially deacetylated acetylphytosphingosine is directly used in the acylation process of the invention, without application of an intermittent recovery and/or drying step.
As salts of sphingoid bases, the HCl or sulphate salts are preferred.
The organic solvent which is employed in the coupling reaction can be any solvent resulting in a sufficient dissolution of the amino alcohol, i.e. a complete dissolution or a dissolution to an extent to keep the amino alcohol sufficiently susceptible to the subsequent reaction with the acid halogenide. Preferably, the organic solvent is a halogenated hydrocarbon, an ester, an ether, a ketone, an alkane, an aromatic hydrocarbon or an aromatic alcohol. More preferably, the organic solvent is a solvent which does not give rise to peroxide formation. Even more preferably, the organic solvent is not a solvent that is miscible with water in all mixing ratios. Even more preferably, the organic solvent is an alkyl ester of formic acid, acetic acid or propionic acid, the alkyl group being a methyl, ethyl, propyl, isopropyl, butyl or isobutyl group. Most preferably, the organic solvent is an alkyl ester of acetic acid, such as ethyl acetate, isopropyl acetate or butyl acetate. The use of alkyl esters of acetic acid has the advantage that these solvents are easily recovered from the process and optionally reused, are cheap and are no peroxide formers.
In the method of the invention, the amino alcohol is suspended in the organic solvent in the additional presence of water. The water can be added either during or after suspending the amino alcohol in the organic solvent, or can originate from the amino alcohol source, or both. The amount of water which may be present in the suspension or solution of the amino alcohol in the organic solvent conveniently may vary from 0.01 to 10 times the volume of the organic solvent. Preferably, the amount of water is 0.1 to 0.5 times the volume of the organic solvent. The upper limit of the amount of water generally will be determined by technical reasons, for instance by the volume of the reaction vessel.
The concentration wherein the amino alcohol is present in the organic solventxe2x80x94water mixture conveniently may amount from 1 to 900 g/l, preferably from 25 to 250 g/l.
In a subsequent step in the process of the invention, the acid halogenide is added under stirring to the amino alcoholxe2x80x94organic solventxe2x80x94water mixture, whereby the acid halogenide may be added as a pure compound or as a solution or suspension in the organic solvent used to suspend the amino alcohol.
In a preferred embodiment of the invention, the acid halogenide is an acid chloride.
The present invention also envisages the option to prepare the acid halogenide in situ by reacting an appropriate organic acid as defined above with a halogenating agent. Suitable halogenating agents are phosphorous trichloride or thionyl chloride.
The acid halogenide typically is added to the amino alcohol organic solventxe2x80x94water mixture in an amount which is a molar equivalent of or which is a slight molar excess to the amount of amino alcohol.
While adding the acid halogenide to the amino alcoholxe2x80x94organic solventxe2x80x94water suspension, the pH of the reaction mixture is kept at a value within a range of 5 to 12, preferably within a range of 6 to 11, by controlled addition of a base. The present invention advantageously allows that pH control is performed using a simple mineral base, such as NaOH. To keep the pH at a stable level during coupling, it is an option to add part of the base as sodium or potassium carbonate.
The temperature during the coupling reaction conveniently is held at a value to allow a thorough stirring of the mixture. The temperature during coupling may also depend on the boiling point of the solvent used. The higher this boiling point, the higher the temperature which may be applied. Typically, for preparation of N-acylated amino alcohols with a low solubility such as certain ceramides, the temperature is kept at a value of 40-80xc2x0 C.
The reaction mixture is stirred for a sufficient time period to allow a substantially complete conversion of the amino alcohol to the N-acylated compound. The extent of conversion conveniently is checked by methods well known in the art, e.g. thin layer chromatography, NMR spectroscopy, HPLC, and the like. Typically, substantially complete conversion is obtained after a time period of about 30 to 60 minutes. In the case that no complete conversion is obtained, additional acid chloride may be added.
In one embodiment of the invention, a halogen substuent present in the R group of the organic acid RCOOH is converted into an oxygen function, subsequent to the coupling stage, in the same organic solvent as used in the coupling stage.
The N-acylated amino alcohol which is produced in the process of the invention is recovered from the organic phase using standard technology. The recovery process generally is a simple process, including washing of the organic phase with water and destining off the water-solvent azeotrope.
Briefly, after completion of the reaction, the temperature of the reaction mixture is increased to a value which is just below the boiling point of the mixture. The water layer is removed and the organic layer optionally is washed with water. The water-solvent azeotrope is distilled off. Optionally, a hot filtration step to remove solid particles and/or an active coal treatment for decolorization may be applied. The product is recovered from the organic phase by cooling and filtration of the crystalline product.
The N-acylated amino alcohol as prepared by the method of the invention preferably is a ceramide, a cerebroside, a retinoyl-sphingoid base, a salicyl-sphingoid base or a short-chain xcex1-hydroxyacyl-sphingoid base. These compounds preferably are applied in cosmetic or dermatological compositions.